Identification of HLA-DRPheβ47 as the susceptibility marker of hypersensitivity to beryllium in individuals lacking the berylliosis-associated supratypic marker HLA-DPGluβ69

BACKGROUND: Susceptibility to beryllium (Be)-hypersensitivity (BH) has been associated with HLA-DP alleles carrying a glutamate at position 69 of the HLA-DP β-chain (HLA-DPGlu69) and with several HLA-DP, -DQ and -DR alleles and polymorphisms. However, no genetic associations have been found between BH affected subjects not carrying the HLA-DPGlu69 susceptibility marker. METHODS: In this report, we re-evaluated an already described patient populations after 7 years of follow-up including new 29 identified BH subjects. An overall population 36 berylliosis patients and 38 Be-sensitization without lung granulomas and 86 Be-exposed controls was analysed to assess the role of the individual HLA-class II polymorphisms associated with BH-susceptibility in HLA-DPGlu69 negative subjects by univariate and multivariate analysis. RESULTS: As previously observed in this population the HLA-DPGlu69 markers was present in higher frequency in berylliosis patients (31 out of 36, 86%) than in Be-sensitized (21 out of 38, 55%, p = 0.008 vs berylliosis) and 41 out of 86 (48%, p < 0.0001 vs berylliosis, p = 0.55 vs Be-sensitized) Be-exposed controls. However, 22 subjects presenting BH did not carry the HLA-DPGlu69 marker. We thus evaluated the contribution of all the HLA-DR, -DP and -DQ polymorphisms in determining BH susceptibility in this subgroup of HLA-Glu69 subjects. In HLA-DPGlu69-negatives a significant association with BH was found for the HLA-DQLeu26, for the HLA-DRB1 locus residues Ser13, Tyr26, His32, Asn37, Phe47 and Arg74 and for the HLA-DRB3 locus clusterized residues Arg11, Tyr26, Asp28, Leu38, Ser60 and Arg74. HLA-DRPhe47 (OR 2.956, p < 0.05) resulting independently associated with BH. Further, Be-stimulated T-cell proliferation in the HLA-DPGlu69-negative subjects (all carrying HLA-DRPhe47) was inhibited by the anti-HLA-DR antibody (range 70–92% inhibition) significantly more than by the anti-HLA-DP antibody (range: 6–29%; p < 0.02 compared to anti-HLA-DR) while it was not affected by the anti-HLA-DQ antibody. CONCLUSION: We conclude that HLA-DPGlu69 is the primary marker of Be-hypersensitivity and HLA-DRPhe47 is associated with BH in Glu69-negative subjects, likely playing a role in Be-presentation and sensitization

A case of pulmonary tuberculosis presenting as diffuse alveolar haemorrhage: is there a role for anticardiolipin antibodies?

<p>Abstract</p> <p>Background</p> <p>Diffuse alveolar haemorrhage (DAH) has been rarely reported in association with pulmonary infections.</p> <p>Case Presentation</p> <p>We report the case of a 43 year old immunocompetent man presenting with dyspnoea, fever and haemoptysis. Chest imaging showed bilateral ground glass opacities. Microbiological and molecular tests were positive for <it>Mycobacterium tuberculosis </it>and treatment with isoniazid, rifampicin, ethambutol and pyrazinamide was successful. In this case the diagnosis of DAH relies on clinical, radiological and endoscopic findings. Routine blood tests documented the presence of anticardiolipin antibodies. In the reported case the diagnostic criteria of antiphospholipid syndrome were not fulfilled.</p> <p>Conclusions</p> <p>The transient presence of anticardiolipin antibodies in association with an unusual clinical presentation of pulmonary tuberculosis is intriguing although a causal relationship cannot be established.</p

Study on the effects of nitrilotriproprionic acid and 4,5-dihydroxy-1,3-benzene disulphonate on the fractionation of beryllium in human serum using graphite furnace atomic absorption spectrometry

<p>Abstract</p> <p>Background</p> <p>Occupational exposure to beryllium may cause Chronic Beryllium Disease (CBD), a lung disorder initiated by an electrostatic interaction with the MHC class II human leukocyte antigen (HLA). Molecular studies have found a significant correlation between the electrostatic potential at the HLA-DP surface and disease susceptibility. CBD can therefore be treated by chelation therapy. In this work, we studied the effect of two complexing agents, nitrilotriproprionic acid (NTP) and 4,5-dihydroxy-1,3-benzene disulphonate (Tiron), on the fractionation of beryllium in human serum analysed by graphite furnace atomic absorption spectrometry (GFAAS).</p> <p>Results</p> <p>We found the average serum beryllium concentration of fourteen non-exposed individuals to be 0.53 (± 0.14) μg l^-1, with 21 (± 3)% of the beryllium mass bound to the low molecular weight fraction (LMW), and 79 (± 3)% bound to the high molecular weight fraction (HMW). The addition of Tiron increased the beryllium mass in the HMW fraction, while NTP was not seen to have any influence on the fractionation of beryllium between the two fractions. NTP was, however, shown to complex 94.5% of the Be mass in the LMW fraction. The beryllium GFAAS detection limit, calculated as three times the standard deviation of 10 replicates of the lowest standard (0.05 μg L^-1), was 6.0 (± 0.2) ng L^-1.</p> <p>Conclusion</p> <p>The concentration of beryllium or its fractionation in human serum was not affected by sex or smoking habit. On average, three quarters of the beryllium in serum were found in the HMW fraction. Of the two ligands tested, only Tiron was effective in mobilising beryllium under physiological conditions, thus increasing the Be content in the HMW fraction.</p

T Regulatory Cells Are Markers of Disease Activity in Multiple Sclerosis Patients

FoxP3+ Treg cells are believed to play a role in the occurrence of autoimmunity and in the determination of clinical recurrences. Contradictory reports are, however, available describing frequency and function of Treg cells during autoimmune diseases. We examined, by both polychromatic flow cytometry, and real-time RT-PCR, several Treg markers in peripheral blood mononuclear cells from patients with multiple sclerosis (MS), an autoimmune disease affecting the central nervous system. We found that Tregs, as defined by CD25, CD39, FoxP3, CTLA4, and GITR expression, were significantly decreased in stable MS patients as compared to healthy donors, but, surprisingly, restored to normal levels during an acute clinical attack. We conclude that Treg cells are not involved in causing clinical relapses, but rather react to inflammation in the attempt to restore homeostasis

RTL551 Treatment of EAE Reduces CD226 and T-bet+ CD4 T Cells in Periphery and Prevents Infiltration of T-bet+ IL-17, IFN-γ Producing T Cells into CNS

Recombinant T cell receptor ligands (RTLs) that target encephalitogenic T-cells can reverse clinical and histological signs of EAE, and are currently in clinical trials for treatment of multiple sclerosis. To evaluate possible regulatory mechanisms, we tested effects of RTL therapy on expression of pathogenic and effector T-cell maturation markers, CD226, T-bet and CD44, by CD4+ Th1 cells early after treatment of MOG-35-55 peptide-induced EAE in C57BL/6 mice. We showed that 1–5 daily injections of RTL551 (two-domain I-Ab covalently linked to MOG-35-55 peptide), but not the control RTL550 (“empty” two-domain I-Ab without a bound peptide) or Vehicle, reduced clinical signs of EAE, prevented trafficking of cells outside the spleen, significantly reduced the frequency of CD226 and T-bet expressing CD4+ T-cells in blood and inhibited expansion of CD44 expressing CD4+ T-cells in blood and spleen. Concomitantly, RTL551 selectively reduced CNS inflammatory lesions, absolute numbers of CNS infiltrating T-bet expressing CD4+ T-cells and IL-17 and IFN-γ secretion by CNS derived MOG-35-55 reactive cells cultured ex vivo. These novel results demonstrate that a major effect of RTL therapy is to attenuate Th1 specific changes in CD4+ T-cells during EAE and prevent expansion of effector T-cells that mediate clinical signs and CNS inflammation in EAE

Erythropoietin: A potent inducer of peripheral immuno/inflammatory modulation in autoimmune EAE

Background: Beneficial effects of short-term erythropoietin (EPO) theraphy have been demonstrated in several animal models of acute neurologic injury, including experimental autoimmune encephalomyelitis(EAE)-the animal model of multiple sclerosis. We have found that EPO treatment substantially reduces the acute clinical paralysis seen EAE mice and this improvements is accompanied by a large reduction in the mononuclear cell infiltration and downregulation of glial MHC class II expression within the inflamed CNS. Other reports have recently indicated that peripherally generated anti-inflammatory CD4 +Foxp3 3 regulatory T cells (Tregs) and the IL17-producing CD4+ T helper cell (Th17) subpopulations play key antagonistic roles in EAE pathogenesis. However, no information regardind the effects of EPO theraphy on the behavior of the general mononuclear-lymphocyte population, Tregs or Th17 cells in EAE has emerged. Methods and Findings: We first determined in vivo that EPO theraphy markedly suppressed MOG specific T cell proliferation and sharply reduced the number of reactive dendritic cells (CD11c positive) in EAE lumph modes during both inductive and later symptomatic phases of MOG 35-55 induced EAE. We then determined the effect in vivo of EPO on numbers of peripheral Treg cells and Th17 cells. We found that EPO treatment modulated immune balance in both the periphery and the inflamed spinal cord by promoting a large expansion in Treg cells, inhibiting Th17 polarization and abrogating proliferation of the antigen presenting dendritic cell population. Finally we utilized tissue culture assays to show that exposure to EPO in vitro similarly downregulated MOG-specific T cell proliferation and also greatly suppressed T cell production of pro-inflammatory cytokines. Conclusions: Taken together, our findings reveal an important new locus whereby EPO induces substantial long-term tissue protection in the host through signalling to several critical subsets of immune cells that reside in the peripheral lymphatic system.published_or_final_versio

A structurally distinct TGF-β mimic from an intestinal helminth parasite potently induces regulatory T cells.

Helminth parasites defy immune exclusion through sophisticated evasion mechanisms, including activation of host immunosuppressive regulatory T (Treg) cells. The mouse parasite Heligmosomoides polygyrus can expand the host Treg population by secreting products that activate TGF-β signalling, but the identity of the active molecule is unknown. Here we identify an H. polygyrus TGF-β mimic (Hp-TGM) that replicates the biological and functional properties of TGF-β, including binding to mammalian TGF-β receptors and inducing mouse and human Foxp3+ Treg cells. Hp-TGM has no homology with mammalian TGF-β or other members of the TGF-β family, but is a member of the complement control protein superfamily. Thus, our data indicate that through convergent evolution, the parasite has acquired a protein with cytokine-like function that is able to exploit an endogenous pathway of immunoregulation in the host

A Pilot Study of IL-2Rα Blockade during Lymphopenia Depletes Regulatory T-cells and Correlates with Enhanced Immunity in Patients with Glioblastoma

Preclinical studies in mice have demonstrated that the prophylactic depletion of immunosuppressive regulatory T-cells (T(Regs)) through targeting the high affinity interleukin-2 (IL-2) receptor (IL-2Rα/CD25) can enhance anti-tumor immunotherapy. However, therapeutic approaches are complicated by the inadvertent inhibition of IL-2Rα expressing anti-tumor effector T-cells.To determine if changes in the cytokine milieu during lymphopenia may engender differential signaling requirements that would enable unarmed anti-IL-2Rα monoclonal antibody (MAbs) to selectively deplete T(Regs) while permitting vaccine-stimulated immune responses.A randomized placebo-controlled pilot study was undertaken to examine the ability of the anti-IL-2Rα MAb daclizumab, given at the time of epidermal growth factor receptor variant III (EGFRvIII) targeted peptide vaccination, to safely and selectively deplete T(Regs) in patients with glioblastoma (GBM) treated with lymphodepleting temozolomide (TMZ).Daclizumab treatment (n = 3) was well-tolerated with no symptoms of autoimmune toxicity and resulted in a significant reduction in the frequency of circulating CD4+Foxp3+ TRegs in comparison to saline controls (n = 3)( p = 0.0464). A significant (p<0.0001) inverse correlation between the frequency of TRegs and the level of EGFRvIII specific humoral responses suggests the depletion of TRegs may be linked to increased vaccine-stimulated humoral immunity. These data suggest this approach deserves further study.ClinicalTrials.gov NCT00626015

Prolonged Survival of Allografts Induced by Mycobacterial Hsp70 Is Dependent on CD4+CD25+ Regulatory T Cells

Background: Heat shock proteins (Hsps) are stress induced proteins with immunomodulatory properties. The Hsp70 of Mycobacterium tuberculosis (TBHsp70) has been shown to have an anti-inflammatory role on rodent autoimmune arthritis models, and the protective effects were demonstrated to be dependent on interleukin-10 (IL-10). We have previously observed that TBHsp70 inhibited maturation of dendritic cells (DCs) and induced IL-10 production by these cells, as well as in synovial fluid cells. Methodology/Principal Findings: We investigated if TBHsp70 could inhibit allograft rejection in two murine allograft systems, a transplanted allogeneic melanoma and a regular skin allograft. In both systems, treatment with TBHsp70 significantly inhibited rejection of the graft, and correlated with regulatory T cells (Tregs) recruitment. This effect was not tumor mediated because injection of TBHsp70 in tumor-free mice induced an increase of Tregs in the draining lymph nodes as well as inhibition of proliferation of lymph node T cells and an increase in IL-10 production. Finally, TBHsp70 inhibited skin allograft acute rejection, and depletion of Tregs using a monoclonal antibody completely abolished this effect. Conclusions/Significance: We present the first evidence for an immunosuppressive role for this protein in a graft rejection system, using an innovative approach - immersion of the graft tissue in TBHsp70 solution instead of protein injection. Also, this is the first study that demonstrates dependence on Treg cells for the immunosuppressive role of TBHsp70. This finding is relevant for the elucidation of the immunomodulatory mechanism of TBHsp70. We propose that this protein can be used not only for chronic inflammatory diseases, but is also useful for organ transplantation management.Pontificia Universidade Catolica do Rio Grande do Sul (PUCRS)Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq)Financiadora de Estudos e Projetos (FINEP

Patterns of Reproductive Isolation in Toads

Understanding the general features of speciation is an important goal in evolutionary biology, and despite significant progress, several unresolved questions remain. We analyzed an extensive comparative dataset consisting of more than 1900 crosses between 92 species of toads to infer patterns of reproductive isolation. This unique dataset provides an opportunity to examine the strength of reproductive isolation, the development and sex ratios of hybrid offspring, patterns of fertility and infertility, and polyploidization in hybrids all in the context of genetic divergence between parental species. We found that the strength of intrinsic postzygotic isolation increases with genetic divergence, but relatively high levels of divergence are necessary before reproductive isolation is complete in toads. Fertilization rates were not correlated to genetic divergence, but hatching success, the number of larvae produced, and the percentage of tadpoles reaching metamorphosis were all inversely related with genetic divergence. Hybrids between species with lower levels of divergence developed to metamorphosis, while hybrids with higher levels of divergence stopped developing in gastrula and larval stages. Sex ratios of hybrid offspring were biased towards males in 70% of crosses and biased towards females in 30% of crosses. Hybrid females from crosses between closely related species were completely fertile, while approximately half (53%) of hybrid males were sterile, with sterility predicted by genetic divergence. The degree of abnormal ploidy in hybrids was positively related to genetic divergence between parental species, but surprisingly, polyploidization had no effect on patterns of asymmetrical inviability. We discuss explanations for these patterns, including the role of Haldane's rule in toads and anurans in general, and suggest mechanisms generating patterns of reproductive isolation in anurans